Flap endonuclease-1 (FEN1) is a 43-kDa metal-dependent nuclear enzyme that exhibits both DNA structure-specific endonuclease activity and 5′ exonuclease activity. Arguably, the most well studied aspect of FEN1 function is its role in the cleavage of Okazaki fragments during DNA replication, Bambara, R. A.; Murante, R. S.; Henricksen, L. A. J. Biological Chemistry 1997, 272, 4647-4650. However, FEN1 also acts to cleave 5′ DNA flaps generated during a variety of other cellular processes including double-strand break repair, Harrington, J. J.; Lieber, M. R. EMBO J. 1994, 13, 1235-1246, homologous recombination, Pont, K. G.; Dawson, R. J.; Carroll, D. EMBO J., 1993, 12, 23-24, and base excision repair (BER), Harrington, J. J.; Lieber, M. R. Genes Dev. 1994, 8, 1334-1335. BER is an important cellular mechanism for the repair of DNA damage caused by alkylating agents, Parikh, S. S.; Mol, C. D.; Hosfield, D. J.; Tainer, J. A. Current Opinion in Structural Biology 1999, 9, 1, 37-47.
The role of FEN1 in BER is clearly exemplified in a recent report that shows nuclease-defective FEN1 results in increased cellular sensitivity to methylmethane sulfonate (MMS), a potent DNA alkylating agent, Shibata, Y.; Nakamura, T. J. Biological Chemistry 2002, 277, 746-754. Sensitization to DNA damaging agents may improve the therapeutic window of classical chemotherapeutics by lowering the minimum effective dose. For a review of chemosensitization: Gesner, T. G.; Harrison, S. D., Annual Reports in Medicinal Chemistry, 2002, 37, 115-124. Recent reports describe several small molecule inhibitors of DNA repair proteins including poly(ADP-ribose) polymerase-1 (PARP) White, A. W.; Almassy, R.; Calvert, A. H.; Curtin, N. J.; Griffin, R. J.; Hostomsky, Z.; Maegley, K.; Newell, D. R.; Srinivasan, S.; Golding, B. T. J. Medicinal Chemistry 2000, 43, 4084-4097, and O6-alkylguanine-DNA alkyltransferase (ATase or MGMT), McElhinney, R. S.; Donnelly, D. J.; McCormick, J. E.; Kelly, J.; Watson, A. J.; Rafferty, J. A.; Elder, R. H.; Middleton, M. R.; Willington, M. A.; McMurry, B. H.; Margison, G. P. J. Medicinal Chemistry, 1998, 41, 5265-5271. These inhibitors are reported to potentiate the activity of various chemotherapeutic agents including temozolomide, Middleton, M. R.; Kelly, J.; Thatcher, N.; Donnelly, D. J.; McElhinney, R. S.; McMurry, B. H.; McCormick, J. E.; Margison, G. P. Int. Journal of Cancer, 2000, 85, 248-252, and topotecan, White, et al. In light of this recent work, we embarked on a strategy to identify selective small-molecule inhibitors of FEN1 for use as chemopotentiating agents. While FEN 1 has a role in DNA replication, the inhibition of the enzyme should not lead to cell death because yeast knockouts are viable, albeit at a slower growth rate than wild-type. Moreau, S.; Morgan, E. A.; Symington, L. S. Genetics, 2001, 159, 1423-1433.
FEN1 is highly homologous to a related endonuclease, xeroderma pigmentosum G (XPG), Gary, R.; Ludwig, D. L.; Cornelius, H. L.; MacInnes, M. A.; Park, J. S. J. Biological Chem., 1997, 272, 24522-24529. XPG is part of a repair pathway that excises DNA containing pyrimidine dimers, a common form of damage caused by exposure to UV light. Defects in XPG are known to cause hypersensitivity to UV light, resulting in light-induced skin lesions and carcinoma, Berneburg, M.; Lehmann, A. R. Adv. Genetics, 2001, 43, 71-102. Therefore, selective inhibition of FEN1 over XPG is a key goal of this program.